Diabetic nephropathy (DN) is the most common cause of end stage kidney disease (ESKD) in the United States. As a result, much effort has been devoted to understanding the mechanisms that promote glomerular damage in diabetic kidney disease. A large body of evidence suggests that glomerular podocytes play a pivotal role in the pathogenesis of DN. A reduced number of podocytes is a characteristic feature of both animals and humans with diabetic kidney disease. Because podocytes are terminally differentiated cells with little potential for proliferation, podocytes that are lost cannot be effectively replaced causing instability of glomerular tuft and promoting disease progression. While the etiology of podocyte loss is likely multifactorial, apoptosis is a prominent feature of diabetic kidney disease and is an important cause of podocyte loss in DN. A key regulator of apoptosis is intracellular calcium. Indeed, a large number of signaling pathways implicated in the pathogenesis of diabetic kidney disease increase intracellular calcium levels. Enhanced intracellular calcium levels activate downstream signaling pathways including the calcium activated phosphatase calcineurin (CN). Our lab has found that receptor systems that increase cytosolic calcium levels and stimulate CN activity promote podocyte apoptosis by mechanisms that are mediated by CN-dependent activation of NFAT (nuclear factor of activated T cells). An important gene target of CN signaling is the transient receptor potential cation channel C6 (TRPC6). These observations appear relevant to DN because: 1. TRPC6 is upregulated in a mouse model of type 1 diabetes (Akita mice), 2. The CN inhibitor FK506 attenuates podocyte apoptosis in Akita mice, and 3. Albuminuria is reduced in Akita mice lacking TRPC6. Based on these observations, we hypothesized that CN plays a key role in DN through mechanisms that involve up-regulation of TRPC6. To investigate this hypothesis, 3 specific aims are proposed. In specific aim #1, we will investigate the dynamic shuttling of NFAT isoforms between the cytosol and nucleus in cultured podocytes, which is dependent on the relative activity of CN phosphatases and NFAT kinases. The goal of these studies is to identify novel strategies for modulating CN- NFAT signaling in glomerular diseases. In specific aim #2, we will determine if the absence of TRPC6 inhibits podocyte apoptosis and reduces glomerular injury in Akita mice lacking TRPC6. In specific aim #3, we will determine if podocyte specific deletion of CN decreases podocyte apoptosis in DN and, in turn, ameliorates kidney injury in Akita mice. The long-term goal of the proposed studies is to identify novel therapeutic targets for the treatment of DN in humans.